Column chromatography provides an excellent means for separating an analyte from other chemical species in a solution. Column chromatography is a separation technique which requires a solvent as the mobile phase and a finely divided solid packing medium as the stationary phase. The molecules of the analyte to be separated interact with the stationary phase but presumably not to the same extent as molecules of other chemical species present in the solution. Thus, when the solution is introduced to the packing medium and is flushed through with eluant, the analyte and various other chemical species present in the original solution distribute themselves between the solvent (the mobile phase) and the stationary phase. The relative time required to pass through the stationary phase varies for the analyte and the other chemical species, thereby providing for separation.
Equally important as an understanding of what is taking place during the column chromatography process is the ability to accurately monitor the process. In other words, ascertaining existing column conditions during separation can provide much useful information such as how best to prepare a column and complete the separation process with the utmost efficiency. State of the art technology provides for the monitoring of several conditions during the column chromatography process. These are, for example, measurement of pH, pressure, conductivity, and UV-absorbing species at inlet and outlet positions of the column.
U.S. Pat. No. 4,165,219 to Huber provides an example of state of the art technology for monitoring a column chromatography process. The Huber patent discloses a device for analyzing solutions utilizing a chromatographic column, a metering device and a detector cell. The detector cell is positioned at the outlet of the chromatography column. While somewhat useful, it must be appreciated that the device in Huber only allows monitoring after a separation is completed. Thus, it fails to provide any specific information about the dynamics of the separation process itself. Such information would be invaluable in understanding the separation process so that a column providing for more efficient separation of a selected analyte may be developed. It should therefore be appreciated that current monitoring systems suffer from significant limitations and shortcomings that need to be overcome.
More specifically, while inlet and outlet monitoring provides information for corrective action downstream or in subsequent batches, the purification system as a whole remains passive. In addition, inlet and outlet monitoring fails to capture or identify many of the important effects that occur within the column itself. These effects include, for example, boundary distortions or band broadening caused by nonequilibrium phenomena at the column inlet and outlet, development of reaction zones among interacting components in the column, and development of gradients and inhomogeneities during the chromatographic process. All of these effects can cooperate to reduce the overall efficiency of the column.
Thus, it is clear that the present technology is inadequate in providing an effective means for monitoring the conditions within the column during the separation process. Such real time monitoring of a selected analyte and other chemical species inside a chromatography column would prove useful in column performance studies, in process or method development, and in process control and validation. Furthermore, a detailed knowledge of conditions inside the column and the flow of analytes and other chemical species of interest, as well as, key impurities, is the first step toward active, intelligent process control.
In an effort to overcome the limitations described above, new techniques are being developed for monitoring conditions within a chromatography column during the separation process. More specifically, optical fibers and metal wires are currently used for transmittal of signals generated by the presence of selected analytes within a column. This has shown to be an effective means for monitoring some conditions within the column chromatography process.
For example, the article titled "Column-Profile Measurements Using Fiber Optic Spectroscopy", published in Soil Science Society of American Journal, Volume 52, 624-7 (1988), discusses the desirability of being able to make measurements to identify flow patterns within a column and making such measurements using fiber optic technology. In addition, the article "Hydrodynamic Studies in Large Diameter Columns", published in Journal of Chromatography, Volume 363, No. 1, 113-23 (1986), discusses using metal wire sensors for studying conditions within the column during the chromatography process.
More particularly, this technology provides for the insertion of an optical fiber or metal wire into a column through an entry port. These entry ports are only large enough to allow for the insertion of a single optical fiber or wire, i.e. one sensor per entry port.
While providing a competent means for in-column monitoring, there are nonetheless still limitations and shortcomings which need to be addressed. For example, with only a single monitoring point established at each port, virtually no flexibility for altering monitoring locations is provided. Specifically, columns with entry ports are specially made and therefore, it would be necessary to reconfigure and construct a new column each time new monitoring locations are desired. In effect only one type of monitoring for a particular chemical species may be completed at each port. This severely limits the useful application of this type of technology.
In addition, state of the art technology, which utilizes optical fibers or metal wires as sensors, further reduces monitoring flexibility by failing to provide any means for effectively adjusting the relative position of the separation zone with respect to the column and entry ports. Thus, it is clear that state-of-the-art approaches are somewhat restricted or limited to monitoring a particular analyte at only certain designated points within the column. Accordingly, the overall value of the real-time information obtained is likewise restricted or limited.